Project Summary/Abstract. Smoking is one of the leading causes of mortality worldwide and is the single greatest, preventable cause of cancer and heart disease. Tobacco use is widespread (1.3 billion smokers worldwide) and current estimates predict that it will contribute to 10 million deaths per year by 2020. The key psychoactive component in tobacco is an alkaloid known as nicotine, which mediates the habit-forming effects of smoking tobacco. Smoking cessation aids on the market consist of pharmacotherapies including nicotine replacement therapy (nicotine patches and gum), bupropion (antidepressant) and varenicline (partial nicotinic receptor agonist). The primary drawback to these currently available treatments is a lack of efficacy; only 20% of patients achieve nicotine abstinence. Clearly, an unmet need exists for a more effective nicotine cessation therapy. In contrast to small-molecule drugs, immunotherapy presents a promising option: nicotine-binding antibodies are administered through active or passive immunization, which sequester nicotine in the blood, thus precluding entry to the brain. Ample preclinical studies have shown that antibodies acting through this mechanism can effectively attenuate nicotine?s addictive effects. Despite the therapeutic potential of anti-nicotine antibodies, no antibodies have been tested in clinical trials as smoking cessation aids. As a possible explanation, previously reported monoclonal antibodies show relatively weak affinity for nicotine, and therefore would not possess sufficient potency for clinical testing. Our goals in this proposal are to produce high affinity anti-nicotine antibodies through state of the art vaccine design. We plan to accomplish our goals of developing an effective antibody through three specific aims: 1. Designing a novel trivalent nicotine conjugate vaccine formulated with the most advanced known adjuvant system 2. Immunize mice with the trivalent nicotine vaccine for generation of antibody-secreting hybridomas, which will be screened and selected based on nicotine affinity 3. In vivo assessment of the selected antibody candidate in rats for mitigating nicotine pharmacology. Completion of the grant aims is anticipated to generate valuable proof of concept data, enabling further development of novel therapeutics for smoking cessation.